title: Evolved Quorum sensing regulator, LsrR, for altered switching functions.
abstract: In order to carry out innovative complex, multistep synthetic biology functions, members of a cell population often must communicate with one another to coordinate processes in a programmed manner. It therefore follows that native microbial communication systems are a conspicuous target for developing engineered populations and networks. Quorum sensing (QS) is a highly conserved mechanism of bacterial cell-cell communication and QS-based synthetic signal transduction pathways represent a new generation of biotechnology toolbox members. Specifically, the E. coli QS master regulator, LsrR, is uniquely positioned to actuate gene expression in response to a QS signal. In order to expand the use of LsrR in synthetic biology, two novel LsrR switches were generated through directed evolution: an "enhanced" repression and derepression eLsrR and a reversed repression/derepression function "activator" aLsrR. Protein modeling and docking studies are presented to gain insight into the QS signal binding to these two evolved proteins and their newly acquired functionality. We demonstrated the use of the aLsrR switch using a coculture system in which a QS signal, produced by one bacterial strain, is used to inhibit gene expression via aLsrR in a different strain. These first ever AI-2 controlled synthetic switches allow gene expression from the lsr promoter to be tuned simultaneously in two distinct cell populations. This work expands the tools available to create engineered microbial populations capable of carrying out complex functions necessary for the development of advanced synthetic products.
authors.affiliation: Fischell Department of Bioengineering, ‡Institute for Bioscience and Biotechnology Research, §Department of Chemical and Biomolecular Engineering, and ∥Department of Chemistry and Biochemistry, University of Maryland , College Park, Maryland 20742 United States .
authors.forename: Bryn L
authors.initials: BL
authors.lastname: Adams
authors.forename: Karen K
authors.initials: KK
authors.lastname: Carter
authors.forename: Min
authors.initials: M
authors.lastname: Guo
authors.forename: Hsuan-Chen
authors.initials: HC
authors.lastname: Wu
authors.forename: Chen-Yu
authors.initials: CY
authors.lastname: Tsao
authors.forename: Herman O
authors.initials: HO
authors.lastname: Sintim
authors.forename: James J
authors.initials: JJ
authors.lastname: Valdes
authors.forename: William E
authors.initials: WE
authors.lastname: Bentley
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D018920:Coculture Techniques; D019020:Directed Molecular Evolution; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation; D053038:Quorum Sensing; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics
chemical_list: D029968:Escherichia coli Proteins; C504601:LsrR protein, E coli; D012097:Repressor Proteins